Background: Spinal cord injury (SCI) can lead to severe motor and sensory dysfunction with high disability and mortality. In recent years, mesenchymal stem cell (MSC)-secreted nano-sized exosomes have shown great potential for promoting functional behavioral recovery following SCI. However, MSCs are usually exposed to normoxia in vitro, which differs greatly from the hypoxic micro-environment in vivo. Thus, the main purpose of this study was to determine whether exosomes derived from MSCs under hypoxia (HExos) exhibit greater effects on functional behavioral recovery than those under normoxia (Exos) following SCI in mice and to seek the underlying mechanism. Methods: Electron microscope, nanoparticle tracking analysis (NTA), and western blot were applied to characterize differences between Exos and HExos group. A SCI model in vivo and a series of in vitro experiments were performed to compare the therapeutic effects between the two groups. Next, a miRNA microarray analysis was performed and a series of rescue experiments were conducted to verify the role of hypoxic exosomal miRNA in SCI. Western blot, luciferase activity, and RNA-ChIP were used to investigate the underlying mechanisms. Results: Our results indicate that HExos promote functional behavioral recovery by shifting microglial polarization from M1 to M2 phenotype in vivo and in vitro. A miRNA array showed miR-216a-5p to be the most enriched in HExos and potentially involved in HExos-mediated microglial polarization. TLR4 was identified as the target downstream gene of miR-216a-5p and the miR-216a-5p/TLR4 axis was confirmed by a series of gain-and loss-offunction experiments. Finally, we found that TLR4/NF-κB/PI3K/AKT signaling cascades may be involved in the modulation of microglial polarization by hypoxic exosomal miR-216a-5p. Conclusion: Hypoxia preconditioning represents a promising and effective approach to optimize the therapeutic actions of MSC-derived exosomes and a combination of MSC-derived exosomes and miRNAs may present a minimally invasive method for treating SCI.

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Spinal cord injury (SCI) can cause severe irreversible motor dysfunction and even death. Neural stem cell (NSC) transplantation can promote functional recovery after acute SCI in experimental animals, but numerous issues, including low-transplanted cell survival rate, cell de-differentiation, and tumor formation need to be resolved before routine clinical application is feasible. Recent studies have shown that transplanted stem cells facilitate regeneration through release of paracrine factors. Small extracellular vesicles (sEVs), the smallest known membrane-bound nanovesicles, are involved in complex intercellular communication systems and are an important vehicle for paracrine delivery of therapeutic agents. However, the application of NSC-derived small extracellular vesicles (NSC-sEVs) to SCI treatment has not been reported. We demonstrate that NSC-sEVs can significantly reduce the extent of SCI, improve functional recovery, and reduce neuronal apoptosis, microglia activation, and neuroinflammation in rats. Furthermore, our study suggests that NSC-sEVs can regulate apoptosis and inflammatory processes by inducing autophagy. In brief, NSC-sEVs increased the expression of the autophagy marker proteins LC3B and beclin-1, and promoted autophagosome formation. Following NSC-sEV infusion, the SCI area was significantly reduced, and the expression levels of the proapoptotic protein Bax, the apoptosis effector cleaved caspase-3, and the pro-inflammatory cytokines TNF-α, IL-1β, and IL-6 were significantly reduced, whereas the expression level of the anti-apoptotic protein Bcl-2 was upregulated. In the presence of the autophagy inhibitor 3MA, however, these inhibitory effects of NSC-sEVs on apoptosis and neuroinflammation were significantly reversed. Our results show for the first time that NSC-sEV treatment has the potential to reduce neuronal apoptosis, inhibit neuroinflammation, and promote functional recovery in SCI model rats at an early stage by promoting autophagy.

The ECRG4 gene was initially identified and cloned in our laboratory from human normal esophageal epithelium (GenBank accession no. AF325503). We revealed the expression of ECRG4 protein was downregulated in 68.5% (89/130) ESCC samples using tissue microarray. The low ECRG4 protein expression was significantly associated with regional lymph node metastasis, primary tumor size, and tumor stage in ESCC (p < 0.05). ECRG4 mRNA expression was downregulated in ESCC due to the hypermethylation in the gene promoter. The treatment with 5-aza-2 0 -deoxycytidine, which is a DNA methyltransferase inhibitor restored ECRG4 mRNA expression in ESCC cells. The result indicated that promoter hypermethylation may be 1 main mechanism leading to the silencing of ECRG4. The high expression of ECRG4 in patients with ESCC was associated with longer survival compared with those with low ECRG4 expression by Kaplan-Meier survival analysis (p < 0.05). ECRG4 protein was an independent prognostic factor for ESCC by multivariable Cox proportional hazards regression analysis (p < 0.05). The restoration of ECRG4 expression in ESCC cells inhibited cell proliferation, colony formation, anchorage-independent growth, cell cycle progression and tumor growth in vivo (p < 0.05). The transfection of ECRG4 gene in ESCC cells inhibited the expression of NF-jB and nuclear translocation, in addition to the expression of COX-2, a NF-jB target gene, was attenuated. Taken together, ECRG4 is a novel candidate tumor suppressor gene in ESCC, and ECRG4 protein is a candidate prognostic marker for ESCC. ' 2009 UICC

Microtubule (MT) kinesin motor proteins orchestrate various cellular processes (e.g. mitosis, motility and organelle transportation) and have been implicated in human carcinogenesis. Kif18A, a plus-end directed MT depolymerase kinesin, regulates MT dynamics, chromosome congression and cell division. In this study, we report that Kif18A is overexpressed in human breast cancers and Kif18A overexpression is associated with tumor grade, metastasis and poor survival. Functional analyses reveal that ectopic overexpression of Kif18A results in cell multinucleation, whereas ablation of Kif18A expression significantly inhibits the proliferative capability of breast cancer cells in vitro and in vivo. Inhibition of Kif18A not only affects the critical mitotic function of Kif18A but also decreases cancer cell migration by stabilizing MTs at leading edges and ultimately induces anoikis of cells with inactivation of the phosphatidylinositol 3-kinase-Akt signaling pathway. Together, our results indicate that Kif18A is involved in human breast carcinogenesis and may serve as a potential therapeutic target for human breast cancer.

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Transplantation of mesenchymal stem cells (MSCs) has been considered an effective therapeutic treatment for a variety of diseases including bone fracture. However, there are associated complications along with MSCs transplantation. There is evidence to show that exosomes (Exos) derived from MSCs exert a similar paracrine function. In addition, repair capabilities of MSCs decline with age. Hence, this study aims to confirm whether the Exos protective function on osteogenic differentiation and fracture healing from aged MSCs was attenuated. This information was used in order to investigate the underlying mechanism. MSCs were successfully isolated and identified from young and aged rats, and Exos were then obtained. Aged-Exos exhibited significantly attenuated effects on MSCs osteogenic differentiation in vitro and facture healing in vivo. Using miRNA array analysis, it was shown that miR-128-3p was markedly upregulated in Aged-Exos. In vitro experiments confirmed that Smad5 is a direct downstream target of miR-128-3p, and was inhibited by overexpressed miR-128-3p. A series gain-and loss-function experiment indicated that miR-128-3P serves a suppressor role in the process of fracture healing. Furthermore, effects caused by miR-128-3P mimic/ inhibitor were reversed by the application of Smad5/siSmad5. Taken together, these results suggest that the therapeutic effects of MSCs-derived Exos may vary according to differential expression of miRNAs. Exosomal miR-128-3P antagomir may act as a promising therapeutic strategy for bone fracture healing, especially for the elderly.

Constraints on models of scalar and vector leptoquarks decaying to a quark and a neutrino at √ s = 13 TeVThe CMS Collaboration * AbstractThe results of a previous search by the CMS Collaboration for squarks and gluinos are reinterpreted to constrain models of leptoquark (LQ) production. The search considers jets in association with a transverse momentum imbalance, using the M T2 variable. The analysis uses proton-proton collision data at √ s = 13 TeV, recorded with the CMS detector at the LHC in 2016 and corresponding to an integrated luminosity of 35.9 fb −1 . Leptoquark pair production is considered with LQ decays to a neutrino and a top, bottom, or light quark. This reinterpretation considers higher mass values than the original CMS search to constrain both scalar and vector LQs. Limits on the cross section for LQ pair production are derived at the 95% confidence level depending on the LQ decay mode. A vector LQ decaying with a 50% branching fraction to tν, and 50% to bτ, has been proposed as part of an explanation of anomalous flavor physics results. In such a model, using only the decays to tν, LQ masses below 1530 GeV are excluded assuming the Yang-Mills case with coupling κ = 1, or 1115 GeV in the minimal coupling case κ = 0, placing the most stringent constraint to date from pair production of vector LQs.Published in Physical Review D as